胺基酸修飾四苯基乙烯自組裝奈米結構與螢光超分子水凝膠之研究

Abstract

近年來，一系列短胜肽序列導入螢光發色團形成的螢光奈米結構，具有發展生物醫材的潛力。本論文以四苯基乙烯( TPE )為基礎連接極短鏈的胜肽分子形成的螢光水凝膠，並探討其自組裝奈米結構與光物理性質。本論文的第一個主題為四苯基乙烯連接不同親疏水性的單一胺基酸TPE-X( X=Gly, Ala, Ser, Asp )，在中性環境下能以最小分子量自組裝形成超分子水膠。不同親疏水性程度的側鏈( R官能基團 )不僅影響自組裝奈米結構的形貌還有凝膠的穩定性。TPE-Gly能自組裝奈米條帶，其他凝膠因子則顯示奈米片狀，而TPE-Ser則可形成穩定度較高的水凝膠。此外，我們也藉由CD、FTIR 與螢光光譜來說明化合物仰賴氫鍵與苯環間的π－π作用力為自組裝的驅動力形成水凝膠。有趣的是，TPE-Gly為唯一沒有對掌中心的分子，但仍能展現卡藤效應，指出其自組裝的方式與其他四者的不同。因此，我們設計一系列的TPE-X-COOH( X=NH( Gly )、O、CH2 )的水凝膠因子來深入調查TPE-Gly的材料性質。2 wt% TPE-CH2-COOH在中性環境下能自組裝奈米纖維形成高穩定的水凝膠，但TPE-O-COOH卻無法。值得一提的是，膠體稀釋後，發現TPE-Gly的奈米條帶會分支出奈米纖維，與同樣稀釋後的TPE-CH2-COOH的奈米纖維，兩者的奈米尺寸是相近的。我們也透過CD、FTIR、XRD與螢光光譜調查TPE-Gly與TPE-CH2-COOH的自組裝差異，推測TPE-Gly的NH為自組裝形成奈米條帶的關鍵。在生物毒性方面，TPE-Gly與TPE-Ala對WS1與3A6細胞皆展現不錯的細胞存活率，而在細胞顯影的表現上，以TPE-Ala為適合作低分子量的生物顯影試劑。 在第二部分的研究主題，我們成功開發出不同色光的四苯基乙烯衍生物，並鏈結FY雙胜肽形成TPE-Blue-FY、TPE-Green-FY與TPE-Yellow-FY的兩親性凝膠因子系統，其能在鹼性與中性的環境下形成穩定水凝膠。透過酸鹼值的調控，我們發現TPE-Blue-FY形成的奈米纖維會有轉變成奈米管柱的現象，使得我們也進一步控制濃度與時間的參數深入了解TPE-Blue-FY的自組裝過程，為第一個以TPE為基礎形成的雙胜肽奈米管柱的化合物。此外，我們也藉由CD、FTIR、螢光光譜分析四苯基乙烯雙胜肽衍生物在鹼性與中性水溶液下的自組裝差異。在生物毒性方面，TPE-Blue-FY、TPE-Green-FY與TPE-Yellow-FY在3A6的細胞培養中，以TPE-Yellow-FY表現出較高的生物相容性，另外，三者衍生物在細胞顯影上皆展現出自身光色的AIE效應，若考量生物毒性，TPE-Yellow-FY會是一不錯的細胞顯影試劑的選擇。

In recent years, a series of short peptides sequences involve fluorophores to form fluorescent nanostructures reveal a great potential in applications of biomaterials. In this study, we explored tetraphenylethylene(-TPE)-based conjugation with ultrashort peptides to form luminescent hydrogels, and investigated its self-assembled nanostructures and photophysical properties. In the first part of this research, we developed TPE/single amino acid conjugateds TPE-X( X=Gly, Ala, Ser, Asp ) that self-assembled into hydrogel with the smallest molecular weight under neutral environment. Side chains ( R group ) with different hydrophobic/hydrophilic degrees were found not only affected the self-assembled nanostructures but also stability of hydrogelation. TPE-Gly could self-assemble into nanobelts, but others formed nanosheets. TPE-Ser could form high stable hydrogel among TPE/single amino acid derivatives. In addition, we demonstrated that these derivatives depended on hydrogen bonding and aromatic π-π stacking interaction as the driving force to form self-assembled hydrogel, as demonstrated by circular dichroism, FTIR and fluorescence spectrum. Interestingly, TPE-Gly was the only achiral molecule, but it could exhibit cotton effect, which its self-assembled in a differently compared to others. Therefore, we designed a series of TPE-X-COOH( X=NH( Gly )、O、CH2 ) hydrogelators to get a deep insight into material properties of TPE-Gly. 2 wt% TPE-CH2-COOH self-assembled nanofibers to form high stable hydrogel under neutral environment, but TPE-O-COOH couldn’t get a gel under similar condition. It was worth to be mentioned that thin fibers were released from nanobelts after hydrogelation of TPE-Gly was diluted, and the fibers were obtained from diluted hydrogelation of TPE-CH2-COOH. Diameters of the both nanofibers were similar. We investigated the self-assembled difference between TPE-Gly and TPE-CH2-COOH through circular dichroism, FTIR , XRD and fluorescence spectrum. We surmised the NH part of TPE-Gly was a key to form nanobelts. In terms of cytotoxicity, TPE-Gly and TPE-Ala showed good cell viability with WS1 and 3A6 cells. In terms of its application in cell imaging, TPE-Ala was showed to stain cells easily with minimal background. In the second part of the research, we successfully developed TPE-based derivatives with different color conjugated with dipeptide FY to form amphiphilic hydrogelators, TPE-Blue-FY, TPE-Green-FY and TPE-Yellow-FY. They could form stable hydrogels under alkaline and neutral environment. We found that self-assembled nanofibers of TPE-Blue-FY turned into nanotubes through adjusting pH value. We also further investigated the self-assembly process of TPE-Blue-FY by controlling concentration and time, demonstrating TPE-Blue-FY was the first one to form TPE-based dipeptide nanotubes. In addition, we analyzed the difference in the self-assembled process between these dipeptide derivatives under base and neutral aqueous solution by circular dichroism, FTIR and fluorescence spectrum. In cytotoxicity, TPE-Yellow-FY possessed good biocompatibility toward 3A6 cells among these peptide derivatives. Furthermore, the three dipeptide derivatives all exhibited AIE property with emission color themselves. Given the promising results in terms of biocompatibility, it could be concluded that TPE-Yellow-FY was a good choice for contrast medium.